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Cooperability
| Project leader |
Ahel Ivan |
| Project co-leader: |
Andreja Mikoč |
| Administering organization: |
Ruđer Bošković Institute
Bijenička 54, 10002 Zagreb
Croatia |
| Partner Institution/Company: |
University of Oxford, Medical Sciences Division, Sir William Dunn School of Pathology |
| Grant type: |
1B |
| Project title: |
Protein ADP-ribosylation in a model prokaryote Streptomyces coelicolor and human |
| Project summary: |
Post-translational modification of proteins plays an important role in mediating protein interactions or modulating protein function. Protein ADP-ribosylation is post-translational modification in which an ADP-ribose moiety from NAD+ is transferred to a target protein. The covalent attachment of one ADP-ribose is catalysed by mono-ADP-ribosyl transferases (mARTs) and leads to mono-ADP-ribosylation. The subsequent transfer of additional ADP-ribose molecules is catalysed by poly-ADP-ribose polymerases (PARPs) and results in the synthesis of poly-ADP-ribose (PAR) polymers (long negatively charged chains of repeating ADP-ribose units). Poly-ADP-ribose metabolism is thought to be restricted to eukaryotes, but it has been recently suggested to exist in a scattering of bacterial species. Protein ADP-ribosylation is a reversible modification and it can be removed by the hydrolytic action of mono-ADP-ribose protein hydrolases (mARHs) and poly-ADP-ribose glycohydrolase (PARG).
This project comprises the investigation of reversible protein ADP-ribosylation and its impact on bacterial metabolism, using the genetic model of Streptomyces coelicolor A3(2), an industrially important bacterium from the genus Streptomyces, which produces over two-thirds of all clinically useful antibiotics of natural origin. Bacteria, and Streptomyces in particular are known to utilize protein ADP-ribosylation to control a variety of important pathways (such as morphological differentiation and antibiotic production), but the proteins involved in this regulation are mostly unknown (transferases and hydrolases). The goal of this project is the identification and characterization of the enzymes controlling reversible protein ADP-ribosylation, as well as the pathways and protein targets for their regulation in Streptomyces. In addition, we will further analyse the physiological consequences of the inactivation of the corresponding genes. Our studies are expected to provide an important understanding on how Streptomyces and potentially other bacteria control their metabolism and antibiotic production. Based on genomic data and our preliminary biochemical experiments, we uncovered the SCO6450 protein in Streptomyces as a potential protein responsible for reversing the protein mono-ADP-ribosylation. The conservation of this protein from bacteria to human (SCO6450 orthologue in human is called MacroD1) should allow the extension of our conclusions and analyses to the human model. We will employ a combination of biochemistry and cell biology to understand the function of MacroD1 in human cells.
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| Hrvatski sažetak: |
Posttranslacijska modifikacija proteina ima važnu ulogu u regulaciji funkcije proteina i interakciji proteina s drugim molekulama. ADP-ribozilacija proteina je posttranslacijska modifikacija kojom se skupina ADP-riboze iz NAD+ prenosi na ciljne proteine. Za proces mono-ADP-ribozilacije odgovorni su enzimi mono-ADP-ribozil transferaze (mART) koji kataliziraju kovalentno vezanje jedne ADP-riboze na ciljni protein. Vezanje dodatnih molekula ADP-riboze kataliziraju poli-ADP-riboza polimeraze (PARP) pri čemu dolazi do sinteze polimera poli-ADP-riboze (PAR) (dugi negativno nabijeni lanci ponavljajućih jedinica ADP-riboze). Iako se smatralo da je metabolizam poli-ADP-riboze vezan isključivo uz eukariote, najnovija istraživanja pokazuju da postoji i kod nekih vrsta bakterija. ADP-ribozilacija proteina je reverzibilna modifikacija i uklanja se hidrolitičkim djelovanjem enzima mono-ADP-riboza protein hidrolaza (mARH) i poli-ADP-riboza glikohidrolaze (PARG).
Predloženi projekt obuhvaća istraživanje reverzibilne ADP-ribozilacije proteina i njenog utjecaja na metabolizam bakterija. Kao genetički model koristit ćemo bakteriju koja pripada rodu Streptomyces, S. coelicolor A3(2). Bakterije ovog roda su od iznimne važnosti jer proizvode preko dvije trećine antibiotika prirodnog porijekla koji imaju kliničku primjenu. Iako bakterije ADP-ribozilacijom proteina kontroliraju važne stanične putove (streptomiceti kontroliraju morfološku diferencijaciju i proizvodnju antibiotika), proteini (transferaze i hidrolaze) uključeni u regulaciju ovog procesa su uglavnom nepoznati. Cilj ovog projekta je identificirati i okarakterizirati enzime koji su odgovorni za proces reverzibilne ADP-ribozilacije proteina kod streptomiceta, kao i stanične putove, te ciljne proteine koje ovi enzimi reguliraju. Nadalje, analizirat ćemo i fiziološke posljedice inaktivacije pojedinih gena uključenih u proces ADP-ribozilacije kod streptomiceta. Očekujemo da će naša istraživanja omogućiti bolje razumijevanje kontrole metabolizma i proizvodnje antibiotika kod streptomiceta (i drugih bakterija). Na temelju genomskih podataka i naših preliminarnih biokemijskih analiza pronašli smo kod S. coelicolor protein SCO6450 koji ima važnu ulogu u procesu mono-ADP-ribozilacije. Očuvanost ovog proteina od bakterija do čovjeka (ortolog SCO6450 kod čovjeka je MacroD1) omogućit će širenje područja našeg istraživanja na ljudski model. Za proučavanje funkcije proteina MacroD1 u ljudskim stanicama koristit ćemo biokemijske metode i metode stanične biologije.
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| Amount requested from UKF: |
1.348.064,28 HRK |
| Amount of matching funding: |
269.612,86 HRK |
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